OHS026
Safe Work Procedure
Faculty/Division
Medicine
School/ Divisional Unit
Inflammatory Diseases Research Unit
Document number
Initial Issue date
Current version
SOMS_IIR_SWP07
January 2010
1.1
Current Version
Issue date
January 2010
Next review date
January 2011
The Writing Safe Work Procedures Guideline (OHS027) should be consulted to assist in the completion of this
form.
Safe Work Procedure Title and basic description
Prepared by Phoebe Phillips 28/7/08
Title: Nitrogen Cavitation of Cells
Description: This task involves lysing cells in a 4°C room using a nitrogen cavitation chamber.
A sample is inserted into the chamber, which is then sealed and nitrogen gas is used to pressurize the chamber. During sample
equilibration at high pressure, nitrogen dissolves into the cells and aqueous buffer. On release of the pressure from the chamber, the
nitrogen rapidly converts back into gas and disrupts the cells as it leaves. The process is carried out in a 4ºC cold room.
Associated risk assessment title and location: 7_RA in the SWP & RA folder, Rm 524, Level 5 Wallace Wurth Building, SoMS
Describe the activity or process
Instructions for Nitrogen Cavitation
Rinse bomb with PBS – make the PBS flow through the outlet tube – you may have to squeeze the tube
to do this.
Close valve at bottom of bomb (VERY important)
Put sample in and screw lid on until tight (Note: This is a lot of turning)
Open valve 4 (ie turn to position
ie. turn anticlockwise)
Attach hose
Turn magnetic stirrer on.
Check Nitrogen valves are closed ie. Twist right.
Open top tap (valve 1) – twist left (anticlockwise – towards you)
Open valve 2 slowly (Note: In tight is off – opposite to a normal
regulator. Turn on by twisting left and loosening)
Pressure in bomb will rise.
Close valve 2 when pressure in bomb is 900psi
Close valve 1
Close bomb valve (valve 4)
Open valve 3 (Note: In-line is Open; across outlet is Off)
Release pressure from housing by opening valve 2
Close both valve 2 and 3
Take hose off
Sit bomb for 1 min
Put tube under outlet hose of bomb.
VERY slowly open bomb valve and collect sample. Note: a lot of foam flows out first
Close valve
Open valve 4 - ie
to release pressure.
Close valve 4 and open chamber.
Rinse bomb and hose with HB+ buffer (~0.5mL) – sit the bomb on the stirrer and then lift up so any
remaining beads are flushed out
Rinse bomb 2X with PBS between samples
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Page 1 of 3
Safe Work Procedure
Uncontrolled document when printed
Date Effective: 01/01/2007
Current Version: 1.2, 15/08/2007
List all resources required including plant,
chemicals, personal protective clothing and
equipment, etc
Personal protective clothing must include gloves, eye goggles/face shield, laboratory coat and extra warm clothing.
Equipment: Nitrogen cavitation chamber, nitrogen cylinder, magnetic stirrer, sample tubes, pipette and tips, MQ-water, kim-wipes
Biological sample(s)
List potential hazards and risk controls including
specific precautions required
1. Cold related illness (shivering, numb hands, loss of fine motor coordination)
Training is essential
Use Safe Work Procedure and read associated risk assessment
Notify a laboratory member you are working in the cold room
Wear warm clothing as well as lab coat and gloves
2. Biological sample spillage (eg Human cell lines)
Wear safety glasses, labcoat and gloves
3. Eye splash
Wear safety glasses, labcoat and gloves
When releasing the sample from the chamber, do this slowly to eliminate splashes and loss of sample.
4. Asphyxiation
Do not close cold room door - leave ajar.
Check cylinder is chained to wall.
Do visual inspections of gas cylinder, regulator and hose.
Check cylinder is turned off.
5. Physical injury.: Cold-burn due to rapid release of nitrogen gas
Wear safety glasses, Labcoat and Gloves
List emergency shutdown instructions
In the event of a gas leak or cylinder de-pressurisation immediately evacuate the area. Notify all other members of the laboratory
and seek assistance from the Lowy Laboratory Manager. This is a major emergency.
Inform the plant custodian and supervisor.
For major emergencies dial x56666
List clean up and waste disposal requirements
1. Cleaning the nitrogen caviation chamber:
Wear gloves during the cleaning procedure. Rinse the chamber and magnetic flea 3 times with MQ-water. Dry with Kimwipes and
return the equipment to their storage place.
Notify the plant custodian if the nitrogen cylinder is low or needs replacing.
2. Cleaning up spills:
Wear safety glasses, labcoat and gloves. Use kimwipes or tissues to wipe up spills then decontaminate surfaces using bleach.
3. Waste disposal:
n/a
List legislation, standards and codes of practice used
in the development of the SWP
AS/NZS 2243.1:2005 Australian/New Zealand Standard™ Safety in laboratories Part 1: Planning and operational aspects
AS/NZS 2243.2:2006 Australian/New Zealand Standard™ Safety in laboratories Part 2: Chemical aspects
AS 2243.6—1990 Australian Standard Safety in laboratories Part 6: Mechanical aspects
AS 2243.3—2000 Australian/New Zealand Standard Safety in laboratories Part 3: Microbiological aspects and containment facilities
NSW OHS Act and Regulation
Supervisory approval, training, and review
Supervisor:
Signature:
Plant custodian: Kat Gaus, CVR, Lowy Building
Signature
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Page 2 of 3
Safe Work Procedure
Uncontrolled document when printed
Date Effective: 01/01/2007
Current Version: 1.2, 15/08/2007
List competency required – qualifications, certificates, licencing, training - eg course or instruction:

UNSW Laboratory Safety Awareness training

UNSW Occupational Health & Safety Awareness training

UNSW Hazardous Substances training

Training in use of the equipment by an authorized user – Hao Kim Lu, Katherine Bryant, Pooli
SWP review date: Jan 2011
Responsibility for SWP review: Katherine Bryant (x52785)
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Page 3 of 3
Safe Work Procedure
Uncontrolled document when printed
Date Effective: 01/01/2007
Current Version: 1.2, 15/08/2007